1. Field of the Invention
The present invention relates in particular to novel chemical compounds, particularly to novel organophosphorus derivatives of indazoles, to compositions containing them, and to the use thereof as medicinal products. More particularly, the invention relates to a series of specific indazoles having anticancer activity, through modulating the activity of proteins, in particular of kinases.
2. Description of the Art
To date, most of the commercial compounds used in chemotherapy are cytotoxic which pose considerable problems of side effects and of tolerance with respect to the patients. These effects could be limited insofar as the medicinal products used act selectively on cancer cells, with the exclusion of normal cells. One of the solutions for limiting the adverse effects of a chemotherapy may therefore consist of the use of medicinal products which act on metabolic pathways or elements constituting these pathways, expressed mainly in cancer cells, and which are not expressed, or expressed very little, in normal cells.
Protein kinases are a family of enzymes which catalyze the phosphorylation of hydroxyl groups of specific residues of proteins such as tyrosine, serine or threonine residues. Such phosphorylations can greatly modify the function of the proteins; thus, protein kinases play an important role in regulating a large variety of cell processes, including in particular metabolism, cell proliferation, cell differentiation, cell migration or cell survival. Among the various cell functions in which the activity of a protein kinase is involved, some processes represent attractive targets for treating cancer-related diseases and also other diseases.
Thus, one of the objects of the present invention is to propose compositions which have anticancer activity, by acting in particular with respect to kinases. Among the kinases for which modulation of the activity is desired, Aurora 2 and Tie2 are preferred.
Many proteins involved in chromosome segregation and spindle assembly have been identified in yeast and drosophila. Disorganization of these proteins results in non-segregation of chromosomes and in monopolar or disorganized spindles. Among these proteins, certain kinases, including Aurora and lpl1, originating respectively from drosophila and from S. cerevisiae, are necessary for chromosome segregation and centrosome separation. A human analog of yeast lpl1 has recently been cloned and characterized by various laboratories. This kinase, called aurora2, STK15 or BTAK, belongs to the serine/threonine kinase family. Bischoff et al. have shown that Aurora2 is oncogenic and is amplified in human colorectal cancers (EMBO J, 1998, 17, 3052-3065). This has also been exemplified in cancers involving epithelial tumors, such as breast cancer.
Among the other kinases on which the products of the invention may act, mention may be made of FAK, KDR, Src, Tie2 and cyclin-dependent kinases (CDKs).
FAK is a cytoplasmic tyrosine kinase which plays an important role in transduction of the signal transmitted by integrins, a family of heterodimeric cell adhesion receptors. FAK and the integrins are located in perimembrane structures called adhesion plaques. It has been shown, in many cell types, that the activation of FAK and also the phosphorylation thereof on tyrosine residues, and in particular the autophosphorylation thereof on tyrosine 397, depend on binding of the integrins to their extracellular ligands, and therefore induced during cell adhesion [Kornberg L, et al. J. Biol. Chem. 267(33): 23439-442. (1992)]. The autophosphorylation of FAK on tyrosine 397 represents a binding site for another tyrosine kinase, Src, via its SH2 domain [Schaller et al. Mol. Cell. Biol. 14:1680-1688. 1994; Xing et al. Mol. Cell. Biol. 5:413-421. 1994]. Src can then phosphorylate FAK on tyrosine 925, thus recruiting the Grb2 adaptor protein and inducing, in certain cells, activation of the ras and MAP kinase pathway involved in the control of cell proliferation [Schlaepfer et al. Nature; 372:786-791. 1994; Schlaepfer et al. Prog. Biophy. Mol. Biol. 71:435-478. 1999; Schlaepfer and Hunter, J. Biol. Chem. 272:13189-13195. 1997]. The activation of FAK can also induce the jun NH2-terminal kinase (JNK) signaling pathway and result in the progression of cells to the G1 phase of the cell cycle [Oktay et al., J. Cell. Biol. 145:1461-1469. 1999]. Phosphatidylinositol-3-OH kinase (Pl3-kinase) also binds to FAK on tyrosine 397 and this interaction could be necessary for the activation of Pl3-kinase [Chen and Guan, Proc. Nat. Acad. Sci. USA. 91:10148-10152. 1994; Ling et al. J. Cell. Biochem. 73:533-544. 1999]. The FAK/Src complex phosphorylates various substrates such as paxillin and p130CAS in fibroblasts [Vuori et al. Mol. Cell. Biol. 16:2606-2613. 1996].
The results of many studies support the hypothesis that FAK inhibitors could be used in the treatment of cancer. Studies have suggested that FAK may play an important role in cell proliferation and/or survival in vitro. For example, in CHO cells, some authors have demonstrated that overexpression of p125FAK results in an acceleration of G1 to S transition, suggesting that p125FAK promotes cell proliferation [Zhao J.-H et al. J. Cell Biol. 143:1997-2008. 1998]. Other authors have shown that tumor cells treated with FAK antisense oligonucleotides lose their adhesion and enter into apoptosis (Xu et al, Cell Growth Differ. 4:413-418. 1996). It has also been demonstrated that FAK promotes cell migration in vitro. Thus, fibroblasts deficient for the expression of FAK (FAK “knockout” mice) exhibit a rounded morphology and deficiencies in cell migration in response to chemotactic signals, and these deficiencies are eliminated by re-expression of FAK [D J. Sieg et al., J. Cell Science. 112:2677-91. 1999]. Overexpression of the C-terminal domain of FAK (FRNK) blocks elongation of adherent cells and reduces cell migration in vitro [Richardson A. and Parsons J. T. Nature. 380:538-540. 1996]. Overexpression of FAK in CHO or COS cells or in human astrocytoma cells promotes cell migration. The involvement of FAK in promoting proliferation and migration of cells in many cell types, in vitro, suggests a potential role for FAK in neoplastic processes. A recent study has effectively demonstrated an increase in tumor cell proliferation in vivo after induction of FAK expression in human astrocytoma cells [Cary L. A. et al. J. Cell Sci. 109:1787-94. 1996; Wang D et al. J. Cell Sci. 113:4221-4230. 2000]. In addition, immunohistochemical studies of human biopsies have demonstrated that FAK is overexpressed in prostate cancers, breast cancers, thyroid cancers, colon cancers, melanomas, brain cancers and lung cancers, the level of expression of FAK being directly correlated with the tumors exhibiting the most aggressive phenotype [Weiner T M, et al. Lancet. 342(8878):1024-1025. 1993; Owens et al. Cancer Research. 55:2752-2755. 1995; Maung K. et al. Oncogene. 18:6824-6828. 1999; Wang D et al. J. Cell Sci. 113:4221-4230. 2000].
KDR (Kinase insert Domain Receptor), also called VEGF-R2 (Vascular Endothelial Growth Factor Receptor 2), is expressed only in endothelial cells. This receptor binds to the angiogenic growth factor VEGF, and thus serves as a mediator for a transduction signal via activation of its intracellular kinase domain. Direct inhibition of the kinase activity of VEGF-R2 makes it possible to reduce the phenomenon of angiogenesis in the presence of exogenous VEGF (Vascular Endothelial Growth Factor: Facteur de croissance vasculaire endothélial) (Strawn et al., Cancer Research, 1996, vol. 56, p. 3540-3545). This process has been demonstrated in particular by means of VEGF-R2 mutants (Millauer et al., Cancer Research, 1996, vol. 56, p.1615-1620). The VEGF-R2 receptor seems to have no function in adults other than that related to the angiogenic activity of VEGF. Consequently, a selective inhibitor of the kinase activity of VEGF-R2 should only show slight toxicity.
In addition to this central role in the dynamic angiogenic process, recent results suggest that VEGF expression contributes to tumor cell survival after chemotherapy and radiotherapy, underlining the potential synergy of KDR inhibitors with other agents (Lee et al. Cancer Research, 2000, vol. 60, p. 5565-5570).
Tie-2 (TEK) is a member of a family of tyrosine kinase receptors, specific for endothelial cells. Tie2 is the first receptor with tyrosine kinase activity for which both the agonist (angiopoietin 1 or Ang1), which stimulates autophosphorylation of the receptor and cell signaling [S. Davis et al (1996) Cell 87, 1161-1169] and the antagonist (angiopoietin 2 or Ang2) [P. C. Maisonpierre et al. (1997) Science 277, 55-60] are known. Angiopoietin 1 can synergize with VEGF in the final stages of neoangiogenesis [Asahara T. Circ. Res.(1998) 233-240]. Knockout experiments and transgenic manipulations of Tie2 expression or of Ang1 expression result in animals which exhibit vascularization deficiencies [D. J. Dumont et al (1994) Genes Dev. 8, 1897-1909 and C. Suri (1996) Cell 87, 1171-1180]. The binding of Ang1 to its receptor results in autophosphorylation of the kinase domain of Tie2, which is essential for neovascularization and for the recruitment and the interaction of the vessels with the pericytes and the smooth muscle cells; these phenomena contribute to the maturation and stability of the newly formed vessels [P. C. Maisonpierre et al (1997) Science 277, 55-60]. Lin et al (1997) J. Clin. Invest. 100, 8: 2072-2078 and Lin P. (1998) PNAS 95, 8829-8834, have shown an inhibition of tumor growth and vascularization, and also a decrease in lung metastases, during adenoviral infections or injections of the extracellular domain of Tie-2 (Tek) in melanoma and breast tumor xenographed models.
Tie2 inhibitors can be used in situations where neovascularization occurs inappropriately (i.e. in diabetic retinopathy, chronic inflammation, psoriasis, Kaposi's sarcoma, chronic neovascularization due to macular degeneration, rheumatoid arthritis, infantile hemangioma and cancers).
The progression of the cell cycle is often controlled by cyclin-dependent kinases (CDK) which are activated by a balance in the cyclin family, which activation ends with the phosphorylation of substrates and, finally, with cell division. In addition, the endogenous CDK inhibitors which are activated (INK4 and KIP/CIP family) negatively regulate CDK activity. Normal cell growth is due to a balance between CDK activators (cyclins) and endogenous CDK inhibitors. In several types of cancers, aberrant expression or activity of several components of the cell cycle has been described.
Cyclin E activates the Cdk2 kinase, which then acts to phosphorylate pRb, resulting in irreversible entry into cell division and transition to the S phase (P L Toogood, Medicinal Research Reviews (2001), 21(6); 487-498), it is also possible, according to these authors, that the CDK2 and CDK3 kinases are necessary for progression in the G1 phase and entry into S phase. During the formation of a complex with cyclin E, they maintain the hyperphosphorylation of pRb so as to aid the progression of the G1 phase to S phase. In the complexes with cyclin A, CDK2 plays a role in the inactivation of E2F and is necessary for realizing the S phase (T D. Davies et al. (2001) Structure 9, 389-3).
The CDK1/cyclin B complex regulates the progression of the cell cycle between the G2 phase and the M phase. Negative regulation of the CDK/cyclin B complex prevents normal cells from entering into S phase before the G2 phase has been correctly and completely effected (K. K. Roy and E. A. Sausville Current Pharmaceutical Design, 2001, 7, 1669-1687.
A level of regulation of CDK activity exists. Cyclin-dependent kinase activators (CAKs) have a positive regulatory action on CDKs. CAK phosphorylates CDKs on the threonine residue so as to render the target enzyme completely active.
The presence of deficiencies in the molecules involved in the cell cycle results in the activation of CDKs and progression of the cycle, thus it seems evident that there is a need to inhibit the activity of the CDK enzymes in order to block cell growth in cancer cells.
All of the references described hereinabove are incorporated herein by reference in their entirety.
The present invention relates to novel organophosphorus derivatives of indazoles. It also relates to the use of organophosphorus derivatives of indazoles modified in the 5-position, as kinase inhibiting agents, and more particularly as anticancer agents. Among these, the invention preferably relates to 5-phosphono- and 5-phosphinoindazoles. It also relates to the use of said derivatives for preparing a medicinal product intended for treating humans.
Among the prior art known to date that describes 5-phosphoindazoles, mention may be made of the patent application published under the number WO93/18008, which is incorporated herein by reference in its entirety. This reference describes derivatives of the formula below:
in which: X=N, CR14 (R14=H, alkyl . . . ); R1=H or halogen; R2=H, NO2, halogen, alkyl . . . ; R3=H, halogen, haloalkyl, haloalkoxy, CN, NH2 . . . ; R4-R6=H, NO2, halo, alkyl, etc., alkylsulfonamido, etc., P(=L)(Q)(M); L=O, S; M, Q=alkoxy, alkyl, (alkyl)namino, OH, H, alkenyloxy, (alkenyl)namino, alkynyloxy, (alkynyl)namino; R7=H, halo, alkyl, NO2; and R8=H, halogen.
Among the compounds disclosed therein, only compounds 147, 161 and 163 are indazoles substituted with a phosphorus-containing group in the 5-position, and are excluded from the present invention as such. On the other hand, these products as medicinal products are part of the present invention. Whereas the compounds disclosed in the aforementioned application have a use in agronomy, i.e., agricultural uses. As mentioned, the compounds of the present invention have a pharmaceutical use.